Trichomonas vaginalis is the causative agent of the most common, non-viral sexually-transmitted infection with ~250 million new cases reported annually in the world and 5-7 million cases in the U.S. In addition to infections of the urogenital tract, trichomoniasis increases the risk of adverse pregnancy outcomes and HIV transmission, and increases the incidence and severity of cervical and prostate cancers. Only two drugs are FDA-approved for the treatment of trichomoniasis, the nitro-heterocyclic compounds metronidazole (Mz) and tinidazole. Oral dosing leads to clinical and microbiological cure in the majority of cases, but treatment failures occur in a significant fraction of patients (up to 17%), with reports of increasing resistance in the U.S. High oral Mz or tinidazole doses in combination with topical treatment of the same drug(s) have been used with success for Mz-refractory vaginal trichomoniasis, whereas topical Mz treatment alone is only modestly effective and so is not recommended. While topical treatment is possible in principle, as shown in murine models, and would be attractive because low systemic absorption is likely to reduce systemic adverse effects compared to oral formulations, current drugs are not sufficiently potent for this preferred administration route. Given the prevalence of trichomoniasis, its association with multiple disease outcomes, and the increase in drug-resistant strains (as well as the potential for more resistance), development of new antimicrobials against trichomoniasis is an urgent need. Our preliminary studies of >1,100 newly synthesized nitro compounds provide compelling evidence that novel nitro drugs can be developed with marked improvements in potency and the ability to completely overcome existing drug resistance. Such next-generation nitro drugs have the distinct advantage that this antimicrobial class, contrary to any other compound class, is already established to be effective and safe against trichomoniasis. The increased potency of our novel compounds should allow development of a topical treatment. Therefore, the project has the overall objective to develop a new lead nitro compound for the topical treatment of Mz-sensitive and Mz-resistant trichomoniasis. Based on our prior data, we have selected several of the most promising structural domains, and will now perform focused structural explorations within these domains. The new derivatives will be tested for activity and potency against Mz-sensitive and Mz- resistant T. vaginalis in vitro, for cytotoxicity and genotoxicity in mammalian cells in vitro, and for efficacy in a murine model of vaginal trichomoniasis in vivo. The comprehensive bioactivity and toxicity data to be generated in this project will be used to select two non-toxic compounds with the best combination of high potency and ability to overcome resistance in vitro, and good efficacy in vivo as lead and back-up for subsequent preclinical development of a novel high-potency compound for the topical treatment of trichomoniasis in phase II of the project.